Abstract Antiretroviral therapy (ART) is critical to improving the health of people living with HIV (PLHIV), to reducing HIV transmission and to maintaining the effectiveness of the current ART programs in resource-limited settings (RLS). However, HIV antiretroviral drug resistance (HIVDR) can hamper global efforts to control the AIDS epidemic and achieve the UNAIDS 90-90-90 targets. Pre-treatment drug resistance (PDR) and on treatment- acquired drug resistance (ADR) are associated with virologic failure (VF) and increased morbidity and mortality. The correlates of PDR and ADR are not fully understood, and the level and breath of HIVDR mutations (DRM) circulating in individuals and populations, especially in RLS is lacking. Much of our understanding of HIVDR genotype-phenotype-outcome correlations have been derived from developed countries where the predominant circulating HIV strain is HIV-1 Group M subtype B. However, in communities with the highest prevalence of HIV-1 infection across the world, PLHIV are infected by non-B subtypes, such as subtypes A, C, D and circulating recombinant forms CRF01_AE and CRF02_AG. HIVDR and the correlates of viral suppression and outcome for these HIV-1 strains has been poorly studied. The success of modern ART regimens in RLS such as TDF-3TC-EFV (TLE) and the planned roll-out of dolutegravir (DTG) based regimens (TLD) could be impeded by emerging evidence of widespread HIVDR. This proposal seeks to expand our understanding of HIVDR and its correlates and consequences in low-and-middle income countries where HIV- 1 non-B subtypes circulate. To accomplish these goals, we propose the following three Specific Aims: 1. Determine the number and breadth of PDR mutations that correlate with virologic failure to TLE or TLD in adults and children infected with HIV-1 subtype A, C, D, CRF01_AE or CRF02_AG. 2. Determine known and novel DRMs at the time of VF in adults and children, including DRM frequencies across cohorts by regimen (TLE and TLD) and by HIV-1 subtype. 3. Determine the correlations between in vitro phenotypic drug resistance testing against genotypic DRMs across HIV-1 non-B subtypes (A, C, D, CRF01_AE and CRF02_AG). To address these aims we will use state-of-the-art and innovative assays to quantify the frequency of DRM in individual?s pre-ART quasispecies and use phenotypic assays to better understand DRM interactions. Our novel studies will determine (1) the risks of specific PDR DRM and minority variants across HIV-1 non-B subtypes for VF; (2) interactions between DRM that determine phenotypic resistance associated with VF; and (3) mutations selected by TLE and TLD regimens at VF. The long-term goal of this proposal is to provide data to enable the best practices for HIV-1 care across a range of resource-limited settings.